A rapid method for quantitation of cell surface IgM by an enzyme-linked immunosorbent assay.
Previous methods for quantitating cell surface immunoglobulins have been relatively tedious or have depended upon use of radioisotopes or specialized equipment (e.g., fluorescence activated cell sorter). We describe a rapid, reproducible method for measuring cell surface Ig on a population of B lymphocytes or lymphoblastoid cell lines by employing an enzyme-linked immunosorbent assay (ELISA) that uses commercially available anti-human Ig-conjugated beads (Immunobeads). Standard curves can be generated that appear to mimic the reaction kinetics of anti-mu with viable cells. Reproducibility of surface mu quantitation depends upon (1) maintaining cell viability during the reaction procedure, and (2) avoiding buffers containing diethanolamine for the alkaline phosphatase reaction. Total Ig in populations of cells can easily be estimated by reacting 10,000 Xg supernatants from lysed cells with anti-Ig beads identical to the standard curve. Combining the methods of surface and total Ig determinations, we were able to analyze quantities of membrane and cytoplasmic mu in populations of lymphoid cells.
D000818:Animals; D001402:B-Lymphocytes / Q000032:analysis / Q000276:immunology*; D002460:Cell Line; D003593:Cytoplasm / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D006041:Goats; D006801:Humans; D007075:Immunoglobulin M / Q000032:analysis*; D007148:Immunoglobulin mu-Chains / Q000032:analysis; D046911:Macromolecular Substances; D014066:Palatine Tonsil / Q000166:cytology / Q000276:immunology; D011947:Receptors, Antigen, B-Cell / Q000032:analysis*; D013154:Spleen / Q000166:cytology / Q000276:immunology; D013601:T-Lymphocytes / Q000032:analysis / Q000276:immunology
D007075:Immunoglobulin M; D007148:Immunoglobulin mu-Chains; D046911:Macromolecular Substances; D011947:Receptors, Antigen, B-Cell